oaioai:CiteSeerX.psu:10.1.1.613.103

Supplemental Experimental Procedures

Abstract

The cDNA of human galectin-3 was amplified from a Caco-2 cDNA library with the primer pair 50-GAATTCTATGGCAGACAATTTTTCGC TCC-30 and 50-GGATCCGGTATCATGGTATATGAAGCACTG-30 and ligated into the unique sites EcoRI/BamHI of the pECFP-C1 or pEYFP-C1 vector (Clontech). The resulting plasmids denoted pCFP/ galectin-3 or pYFP/galectin-3 were confirmed by sequencing. For the generation of prab11-CFP, the cDNA sequence of GFP was substituted by CFP in the vector prab11-GFP [S1]. Vesicle Preparation, Proteinase K Treatment, and TCA Precipitation Post-TGN vesicles of MDCK-SI-YFP and MDCK-LPHmyc cells were isolated as described previously [S2]. For the enrichment of post-TGN vesicles, the cells were incubated in the presence of cyclohex-imide (100 mg/ml) for 4 hr at 20ºC to accumulate newly synthesized material in the TGN, followed by TGN release at 37ºC for definit

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oaioai:CiteSeerX.psu:10.1.1.613.103Last time updated on 10/29/2017

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